NtSKP1基因的反义载体构建及转基因烟草的产生  

Construction the Antisense Expression Vector of NtSKP1 and Production of Transgenic Tobacco

作　　者：张付云[1] 李伟[1] 张瑛[1] 孙秋颖[1] 李研[1] 杜昱光[2] 

机构地区：[1]大连水产学院,大连116023 [2]中国科学院大连化学物理研究所辽宁省碳水化合物研究重点实验室,大连116023

出　　处：《生物技术通报》2010年第4期141-145,共5页Biotechnology Bulletin

基　　金：国家高技术研究发展计划"863"项目(2006AA10A213);国家高技术研究发展计划"863"项目(2007AA091601);中国科学院知识创新工程重要方向项目(KSCX2-YW-G-041;KSCX2-YW-N-007);辽宁省自然科学基金项目(20082153)

摘　　要：根据枯斑三生烟SKP1基因(NtSKP1)的序列,设计一对分别含有特定酶切位点的特异引物,以重组质粒pMD18-SKP1为模板,扩增目的基因(约473bp)片段。将反向目的片段插入中间载体pHANNIBAL的内含子右侧,再经NotⅠ酶切回收约3443bp的目的片段,插入到双元载体质粒pART27中,成功构建了含NtSKP1基因片段反向序列的植物表达载体pART27-skp1a,其转录产物能减弱目的基因的表达。将pART27-skp1a质粒导人根癌农杆菌LBA4404中并转化烟草叶片细胞,经选择分化培养,获得转基因烟草。According to the sequence of SKP1 from Nicotiana tabacum var. Samsun NN （NtSKP1）,one pair of specific primers containing special restriction enzyme sites were designed. The target reverse fragments（473 bp）were amplified and inserted into right side of intron in intermediated vector pHANNIBAL. Then the target fragment about 3 443 bp was cut by NotⅠ from recombinant intermediated vector,and then cloned into binary plasmid pART27. Plant expression plasmid pART27-skp1a,with reverse DNA fragment of NtSKP1 was successfully constructed. And pART27-skp1a was introduced into Agrobacterium tumefaciens LBA4404 and then transformed tobacco leaves.The transcriptional products of the foreign fragment probably decrease the expression of target gene. The transgenic plants of tobacco were obtained.

关 键 词：NtSKP1基因 反义表达载体的构建 转基因烟草 

分 类 号：Q943.2[生物学—植物学]